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In the isolated perfused liver system high concentrations of phenformin (0.93--1.24 mM) were required to reduce the greater than two-fold elevated rate of gluconeogenesis from L-[U-14C]lactate in acutely alloxan diabetic (48-hour) and chronically alloxan diabetic (7-day) rat livers to the slower rate of normal fed livers. At these phenformin concentrations, other hepatic functions such as substrate uptake and 14CO2 production were also inhibited. The livers were also in a very reduced state under these conditions as indicated by the elevated ratios of the redox couples lactate/pyruvate and 3-hydroxybutyrate/acetoacetic acid. The results are interpreted to indicate that if phenformin functions as an antidiabetic (hypoglycemic) agent by inhibiting hepatic gluconeogenesis to normal levels, it is also generally toxic to the liver under such conditions. The results are discussed in relation to current hypotheses of the mechanism of action of phenformin and to phenformin-associated lactic acidosis.

Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of detergent solubilized cytochrome P-450 fractions from normal and diabetic female rat liver microsomes revealed the induction of a 52,000 molecular weight microsomal hemeprotein in diabetic liver. Two major P-450 hemeproteins (DB IIA and DB IIB) were subsequently isolated from solubilized diabetic female rat hepatic P-450 fractions, while normal rat liver yielded only one major microsomal P-450, N IIA. When examined in a reconstituted drug metabolizing system, the aniline hydroxylation rate catalyzed by DB IIB (52,000 molecular weight) was 157% and 78% greater than the rates catalyzed by N IIA and DB IIA, respectively. The rates of ethylmorphine metabolism catalyzed by N IIA and DB IIB were similar; however, the rate of ethylmorphine metabolism catalyzed by DB IIA (54,000 molecular weight) was approximately 42% greater than the rates catalyzed by either N IIA or DB IIB. These results are compared to those previously obtained with P-450s isolated from diabetic male rat liver and indicate that diabetes induces P-450s with specific catalytic activities which can account for both the sex-dependent and sex-independent alterations in drug metabolism observed in diabetic rat liver.

Previous studies have shown that diabetes induces a unique 52,000 molecular weight cytochrome P-450 in rat hepatic microsomes. In the present study, the catalytic properties of the two major purified diabetic P-450s were contrasted to those of the major normal purified P-450 in a reconstituted drug metabolizing system. the greatest rate of aniline hydroxylation was catalyzed by the 52,000 molecular weight diabetic P-450 (7-fold greater than normal). This same diabetic P-450 also catalyzed a slower rate of ethylmorphine N-demethylation compared to the major normal and the other diabetic P-450. These results indicate that the catalytic properties of this diabetes-dependent P-450 are responsible for the substrate-specific alterations in drug metabolism observed in both liver microsomes and isolated perfused liver from diabetic rats.

Sodium dodecyl sulfate-polyacrylamide gel electrophoresis was used to determine the effect of diabetes on the cytochrome P-450-region proteins of rat liver microsomes. Compared to normal microsomes, diabetic microsomes from both male and female rats exhibited distinct increases as well as decreases in the hemeproteins of this region. These results support the concept that the substrate specific changes in drug metabolism observed in diabetic rats may be related to specific alterations in the hepatic cytochrome P-450 hemeprotein population.

The demethylation of aminopyrine was decreased while the demethylation of p-chloro-N-methylaniline was increased in both alloxan and streptozotocin diabetic isolated perfused male rat livers. Since similar effects of diabetes were observed with traditional microsomal preparations, these observations indicate that the effects of diabetes on drug metabolism observed in isolated microsomal preparations from male rat are an accurate reflection of conditions in the intact organ. These results also clearly indicate that diabetes in the male rat affects at least two different rate-limiting steps in the hepatic drug metabolism process.